Costimulatory B7 molecules (B7-1 or CD80; and B7-2 or CD86) are known to bind to T cell costimulatory receptors CD28 and CTLA4. Engagement of CD28 is know to transduce signals in T cells that play a critical role in T cell activation. The meachanism of signal transduction mediating T cell activation and costimulation has been studied. TCR mediated activation of T cells to proliferation and IL2 secretion was enhanced in mice deficient in cbl-b, suggesting a negative regulatory role for cbl-b under these conditions. Cbl-b deletion in fact reversed the inability of CD28-deficient mice to generate T cell-dependent Ig class switched primary and secondary antibody responses. Surprisingly, however, cbl-b deletion did not reverse the functional defect of B7 (CD80/CD86) deficient mice, indicating an unexpected difference bewtween the requirement for CD28 and for CD28 ligand B7 molecules. Moreover, cbl-b inactivation substantially reversed the immune response defect in mice deficient in both CD28 and B7. Overall, these results suggest the unanticipated conclusion that the expression of CD28 has a strong negative regulatory effect on immune response in the absence of its B7 ligands. The mechanism underlying these effects is under study employing CD28 mutants with deletion or inactivation of intracellular or extracellular domains.. Additional studies have been initiated to study the molecular pathways of T cell activation and costimulatory signaling. The relationship between members of the cbl family and another critical adapter molecule, SLP-76, are being studied using additional strategies of genetically manipulated expresssion. Experiments have indicated that inactivation of c-cbl (but not cbl-b) completely reverses the partial lethal phenotype caused by SLP-76 deficiency. In addition, c-cbl inactivation partially reverses the defect in T cell development caused by deficiency in SLP-76 or LAT, resulting in fact in a marked hyperplasia of peripheral CD4 T cells. These findings indicate an unanticipated SLP-76 (and LAT) independent signaling pathway that is facilitated by c-cbl inactivation. The biochemical basis for these effects is under study. The function of B7 in T-dependent B cell activation is being analyzed using B cells from mice that are genetically engineered to be deficient in B7 expression or to express mutated or truncated B7 products. Studies in these mice and in radiation chimeras generated from these mice will determine whether expression of B7 and signal transduction through the B7 cytoplasmic tail are required for T-dependent antibody responses, Ig class switching, and generation of memory B cells.